1. Field of the Invention
The counting of particles in fluid suspensions by fluorescent emission has a wide range of applicability in immunoassay techniques and in the characterization of biological materials in general. Known methods, however, require specially designed orifices, flow conduits, or sensing zones, or complex computational techniques to differentiate the particles of interest from extraneous or undesired components in the sample.
There is thus a need for an inexpensive yet accurate technique which provides a direct indication of particle presence, concentration and/or size, particularly where particles are being detected with low signal to noise ratio.
2. Description of the Prior Art
The use of flow cytometers involving the carefully controlled flow of a cell suspension through a narrow flow channel is described in Miller, et al., "Usage of the Flow Cytometer-Cell Sorter," Journal of Immunological Methods, 47, 13-24 (1981); Hoffman, et al., "Immunofluorescent Analysis of Blood Cells by Flow Cytometry," Int. J. Immunopharmac., 3(3), 249-254 (1981); a general review of flow cytometry is found in Flow Cytometry and Sorting, M. R. Melamed, P. E. Mullaney and M. L. Mendelsohn, Eds., J. Wiley & Sons, N.Y., N.Y. 1979; Hansen, et al., U.S. Pat. No. 4,284,355, issued Aug. 18, 1981; Hansen, et al., U.S. Pat. No. 4,284,412, issued Aug. 18, 1981; Auer, et al., U.S. Pat. No. 4,284,924, issued Aug. 4, 1981; and Stevens, U.S. Pat. No. 3,275,834, issued Sept. 27, 1966.
The use of laser beams and slits to differentiate particles based on their relative size by the correlation of fluorescence fluctuations in a relatively large sample volume is described in: Briggs, et al., "Homogeneous Fluorescent Immunoassay," Science, 212, 1266-1267 (1981) and Nicoli, et al., "Fluorescence Immunoassay Based on Long Time Correlations of Number Fluctuations," Proc. Natl. Acad. Sci. U.S.A., 77(8), 4904-4908 (1980).
U.S. Pat. No. 4,421,860 (Elings, et al.) describes a homogeneous fluoroimmunoassay involving autocorrelation processing of optically sensed signals. U.S. Pat. No. 4,407,964 (Elings, et al.) discloses a homogeneous fluoroimmunoassay involving sensing radiation for forward and back directions.
An immunological reagent and radioimmunoassay are disclosed by Dreyer in U.S. Pat. No. 3,853,987.